Substituted nitroimidazolyl thiadiazoles and oxadiazoles as antibacterial agents and growth promoting compounds

ABSTRACT

The use of substituted nitroimidazolyl-thiadiazoles and oxadiazoles are described along with methods of administration of the same. These compounds are active in enhancing the growth rate of warm-blooded animals and in controlling the growth of pathogenic microorganisms such as bacteria.

United States Patent Berkelhammer et a1. Sept. 9, 1975 SUBSTITUTED NITROIMIDAZOLYL [52] US. Cl. 424/250; 424/248; 424/267; THIADIAZOLES AND OXADIAZOLES AS 424/270; 424/272 ANTIBACTERIAL AGE A GROWTH [51] Int. Cl. A01N 9/22; A6114 31/495 of Search [75] Inventors: Gerald Berkelhammer, Princeton; [56] References Cited I h NJ. Goro Asato T1tusv1lle bot of UNITED STATES PATENTS [73] Assign: America" Cyanamid 3,025,303 3/1962 Ifversen et a1. 424 270 Stamford, Conn- 3,452,035 6/1969 Berkelhammer et a1. 424 272 [22] Filed: Apr. 25, 1974 Primary Examiner--V. D. Turner [2H Appl' 463951 Attorney, Agent, or FirmJack W. Richards Related US. Application Data [60] Division Of Ser. NO. 199,005, Nov. 15, 1971, Pat. NO. [57] ABSTRACT 3,830,924, which is a continuation-in-part of Ser. No. 17,977, March 9, 1970, abandoned, which is a continuation-in-part of Ser. No. 814,205, April 7, 1969, abandoned, which is a continuation-in-part of Ser. No. 659,596, Aug. 10, 1967, Pat. No. 3,452,035, which is a continuation-in-part of Ser. No. 604,158, Dec. 23, 1966, abandoned.

The use of substituted nitroimidazolyl-thiadiazoles and oxadiazoles are described along with methods of administration of the same. These compounds are active in enhancing the growth rate of warm-b1ooded animals and in controlling the growth of pathogenic microorganisms such as bacteria.

1 Claim, N0 Drawings SUBSTITUTED NITROIMIDAZOLYL THIADIAZOLES AND OXADIAZOLES AS ANTIBACTERIAL AGENTS AND GROWTH PROMOTINO COMPOUNDS CROSS-REFERENCE TO RELATED APPLICATIONS The present invention is a divisional of our application Ser. No. 199,005, filed Nov. 15, 1971, now US. Pat. No. 3,830,924, which in turn is a continuation-inpart of our application Ser. No. 17,977, filed Mar. 9, 1970, now abandoned, which is a continuation-in-part of our application Ser. No. 814,205, filed Apr. 7, 1969, now abandoned, which is a continuationin-part of our application Ser. No. 659,596, filed Aug. 10, 1967, now US. Pat. No. 3,452,035, which in turn is a continuation-in-part of our application Ser. No. 604,158, filed Dec. 23, 1966, now abandoned.

BACKGROUND OF THE INVENTION The present invention describes the use and method of administration of compounds which are both highly effective at relatively low concentrations against a broad spectrum of microorganisms, and additionally provide a relatively satisfactory margin of safety. The compositions containing substituted imidazolyl compounds of the present invention are at least 5 to times more active than certain distantly related imidazolyl compounds of theprior art, providing effectiveness at relatively low concentrations, as well as satisfactory margins of safety.

Imidazolyl compounds, in which the heterocyclic rings are joined by a methyleneamine bridge, have been prepared and found to have some antibacterial activity; however, such compounds have not been entirely satisfactory for these purposes. The concentration at which such compounds are active is generally much higher than the desirable level; therefore, said compounds do not provide a satisfactory margin of safety. Illustrative of said compounds which have been prepared and found to have such activity are, for example, 3-[(lmethyl-S-nitro-Z-imidazolylmethylene )amino1-2- oxazolidinone and 1-[(1-methyl-5-nitro-Z-imidazolylmethylene)amino]-2-imidazolidinone.

SUMMARY OF THE INVENTION The subject matter of the present invention relates to compositions containing as the active component 1- substituted-5-nitro-2-imidazo1yl compounds and an edible carrier. More particularly, the invention relates to compositions containing compounds having the following general formula:

wherein R is selected from the group consisting of lower alkyl, hydroxy lower alkyl, lower alkanoyloxy lower alkyl and benzyl; X is selected from the group consisting of oxygen and sulfur; and R, and R are selected from the group consisting of hydrogen, alkyl of 1 2 to 8 carbon atoms, hydroxy lower alkyl, lower alkoxy lower alkyl, cyclohexyl, formyl, lower alkanoyl, monochlorolower allkanoyl, dichlorolower alkanoyl, morpholino lower alkanoyl, lower alkyl aminolower alkyl;

taken together is selected from the group consisting of -N=CHN (lower a1kyl) piperazine, N-lower alkylpiperazine, N-hydroxy lower alkylpiperazine, N- pyridylpiperazine, N-thiazolylpiperazine, N-lower alkoxycarbonyl piperazine, piperidino and dilower alkylamino lower alkylpiperpidino; and physiologically acceptable salts thereof. These salts can be, for example, the hydrochloride hydrobromide, hydroiodide, sulfate, phosphate, etc. The term lower alkyl" as employed in the instant specification and claims is intended to include either straight or branched chain having from 1 to 4 carbon atoms. The invention relates to a method of controlling the growth of bacteria in a warm-blooded animal host which comprises administering to the animal therapeutically effective quantities of the above compounds and thereby alleviating diseases, as well as compositions of matter comprising said compounds and an edible carrier, In addition, methods are described employing the above compounds to promote the growth rate in warm-blooded animals.

PREPARATION OF THE SUBSTITUTED IMIDAZOLYLS OF THE PRESENT INVENTION In many instances the substituted imidazolyl compounds comprising the active component of the present compositions are prepared by oxidative cyclization of various thiosemicarbazones and semicarbazones of 1- substituted-S-nitro-2-imidazolecarboxaldehyde. Suitable oxidizing agents for such cyclization of the thiosemicarbazone to aminothiadiazoles include a wide variety of ferric salts such as ferric ammonium sulfate, ferric chloride, ferric nitrate, ferric acetate, sodium ferricyanide, sodium ferric oxalate, potassium ferric sulfate, and the like. The cyclizations of the semicarbazones to aminooxadiazoles are generally carried out with such agents as sodium hypobromite, sodium hypoiodite, and bromine with sodium acetate. The reactions are generally carried out at an elevated temperature between 50 and 150C, depending on the particular moiety being prepared.

A number of the 2'(2-substituted amino-5- thiadiazolyl)-l-substituted-5-nitroimidazo1es can be prepared by the reaction of 2-(2halo-5-thiadiazolyl)- l-substituted-5-nitroimidazoles with the appropriate primary or secondary amines in an organic solvent, either in the presence of excess amine or with the use of other acid acceptors, as for example, aqueous sodium bicarbonate solution, usually at between 25 and C.

In the preparation of active components having the above general formula wherein R represents a hydroxy lower alkyl group, it is often practical to first synthesize the corresponding ester, as for example, a compound of the formula:

LUK? wherein R is lower alkanoyloxy lower alkyl and X and R are as defined hereinabove, and then treat the thus formed ester with a strong mineral acid followed by pH adjustment to above about pH 7, thereby yielding the desired product. In many cases, it is found that the conditions used for cyclization 1-( Z-lower alkanoyloxy lower alkyl)--nitro-2-imidazole carboxaldehyde thiosemicarbazones are sufficiently acidic to give rise directly to the 2-(2-amino-5-thiadiazolyl)-l-(hydroxy lower alkyl )-5-nitroimidazoles.

Where it is desired to prepare the 2-(2-formamido-5- thiadiazolyl) l-substitutednitroimidazole, or the 2-(2- acylamino-or 2-halocylamino-S-thiadiazolyl)-1-substituted-5-nitroimidazole, it is practical to first synthesize the 2-(2-amino-5-thiadiazolyl)-l-substituted-S- nitroimidazole, and then treat the product with formic acid, in the case of the preparation of the 2-(2-formamido-S-thiadiazolyl l -substituted-5 nitroimidazole, or with an anhydride of the formula (lower alkanoyD- or (halo lower alkanoyl) or with the appropriate acid chloride in the instance of the preparation of the 2-( 2-acylamino or 2-halocylamino-5- thiadiazolyl l -substituted-5 -nitroimidazole. Similar reactions can be carried out to give the corresponding oxadiazolyl compounds. These reactions are usually carried out at an elevated temperature, particularly at temperatures between 50 and 150C.

The 2-(2-amino-5-thiadiazolyl)- 1 -lower alkanoyloxy lower alkyl-5-nitroimidazoles and the corresponding oxadiazolyl compounds, active compounds of the present compositions, can be made by esterifying the 2-( 2- amino-S-thiadiazolyl or 5-oxadiazolyl)-l-hydroxy lower alkyl-S-nitroimidazoles by heating with an aliphatic acid in the presence of a mineral acid catalyst. If, instead, an aliphatic acid anhydride is employed, the products are 2-(2-lower aIkanoylamino-S-thiadiazolyl or 5-oxadiazolyl)-l-lower alkanoyl lower alkyl-5 nitroimidazoles. In the latter case, a catalyst is usually not necessary. In both instances, reaction temperatures of from 50 to 150C. are frequently employed.

For the preparation of N,N-dilower alkyl-N-[5-(lsubstituted-5-nitro-2-imidazolyl )thiadiazol-2-yl]for mamidines, products of the reaction of N,N-dilower alkyl formamides with phosgene, phosphorus oxychloride, or thionyl chloride are reacted with 2-(2-amino-5- thiadiazolyl)-l-substituted-5-nitroimidazoles in an organic solvent, as for example, in an excess of the N,N- dilower alkyl formamide, usually at the ambient temperature.

The active components of the compositions of the instant invention are highly effective in controlling infections of pathogenic and other microorganisms such as Escherichia coli, Salmonella gallinarum, Salmonella clwteraesirs Var Kunzendorf, Proteus mirabilis, Slap/zylococcus aureus strain Rose or strain Smith, Salmonella typlwsa, Klehsiella AD, aerobacter aerogenes, Streptococcus pyogenes, Pasleurella mulmcida, etc. Tests showing the use ofa compound of this invention against bacteria are described in Antimicrobial Agents and Chemotherapy-I968" page 534 etc., in a warmblooded animal host; said active components may be administered to the warm-blooded animals, such as' goats, sheep, cattle, hogs, chickens, etc., in admixture with their feed or drinking water. Furthermore, the compositions may be administered in the form of tablets, pills, capsules or the like, or parenterally by injection either intramuseularly or subcutaneously. The

concentration employed in feed or water may be in the range of from 5 to 1,000 parts per million, preferably 15 to 500 parts per million; the most preferred concentration being about 50 to 200 parts per million. The above compositions have demonstrated effectiveness against Salmonella infections when the active component was administered in as little as 0.025% concentration in the diet of chicks and mice. In addition, some of the active components provide control of Escherichia coli infections in chicks when administered at about 40 mg. per kg. of body weight in a single oral dose and are highly effective against a number of other bacterial infectionss, both gram-positive and gram-negative.

The active components of this invention can be used with pharmaceutically acceptable edible carriers in compositions such as tablets (containing 1 mg. to 100 mg. per kilogram of body weight), wherein the principal active ingredient is mixed with conventional tableting ingredients such as corn starch, lactose, sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalcium phosphate, gums, and fractionally similar materials as pharmaceutical diluents or carriers. The tablets or pills of the novel compositions can be laminated or otherwise compounded to provide a dosage form affording the advantage of prolonged or delayed action of predetermined successive action of the enclosed medication. For example, the tablet or pill can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former. The two components can be separated by an enteric layer which serves to resist disintegration in the stomach, and permits the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymic acids or mixtures of polymeric acids with such materials as shellac, shellac and cetyl alcohol, cellulose acetate and the like. A particularly advantageous enteric coating comprises a styrene maleic acid copolymer together with known materials contributing to the enteric properties of the coating. The term dosage form as described herein refers to physically discrete units suitable as unitary dosage for warm-blooded animal subjects, each unit containing a predetermined quan tity of active material calculated to produce the desired therapeutic effect in association with the required pharmaceutical diluent, carrier or vehicle. Examples of suitable oral dosage forms in accord with this invention are tablets, capsules, pills, powder packets, granules, wafers, cachets, teaspoonfuls, dropperfuls, ampules, vials, segregated multiples of any of the foregoing and other forms as herein described.

The compounds of this invention have been found to promote the growth rate of warmblooded animals, such as goats, sheep, cattle, hogs, chickens, etc. Said compounds may be administered in a manner similar to their introduction as antibacterial agents, such as orally or parenterally.

DETAILED DESCRIPTION The following examples are provided for illustrative purposes and may include particular features of the invention; however, the examples should not be construed as limiting the invention, many variations of which are possible without departing from the'spirit or scope thereof. Parts are by weight unless otherwise in dicated.

EXAMPLE 1,

Preparation of 2-( 2-Amino-5-Thiadiazolyl 1 -methyl-5- nitroimidazole A mixture of 4.71 gm., 0.03 mole, of l-methyl-2- hydroxymethyl-S-nitroimidazole, and 13.3 gm., 0.03 mole, of lead tetraacetate in 200 ml. of benzene, is refluxed while stirring magnetically for about 18 hours, cooled and filtered. The filtrate is washed with 50 ml. of saturated sodium carbonate solution. The organic phase is then separated, and the aqueous phase extracted twice with 30 ml. of chloroform. The combined organic phase is then dried over magnesium sulfate. After filtering and evaporating the organic phase to dryness, the filtrate gives 4.2 gm. of pale yellow 1- methyl-5-nitro 2-imidazolecarboxaldehyde, which is dissolved in 25 ml. of hot ethanol, then added to 2.5 gm. of thiosemicarbazide in 20 ml. of boiling ethanol containing two drops of concentrated hydrochloric acid. The mixture is then boiled for a few minutes with stirring, cooled and bright yellow crystals of the thiosemicarbazone of the above aldehyde are collected. The process yields 5.3 gm., which constitutes a 77.4% overall yield, said material having a melting point of 227C. (decomposition).

To 25 ml. of hot water containing 5.7 gm., of ferric ammonium sulfate dodecahydrate, 2.68 gm. of the above thiosemicarbazone is added, and the mixture is stirred magnetically in a boiling water bath. After 1 hour, an additional 75 ml. of hot water containing 17.1 gm. of ferric ammonium sulfate dodecahydrate is added to the above mixture. The mixture is then heated for approximately 3 hours in a boiling water bath, and filtered while still hot, yielding orange brown crystals which are washed thoroughly with hot'water. The yield is 2.7 gm., having a melting point of from 259 to 260C. (decomposition). This product is dissolved in about 20 ml. of hot dimethylformamide, filtered and the warm filtrate poured on ice. The precipitated product is washed thoroughly first with water, and then cold acetone, giving a yellow product which is dried in vacuo at 100C. for several hours. The purified product weighs 1.55 gm., and has a melting point of from 268 to 270C. (decomposition). The product is then submitted to analysis; calculating from carbon, hydrogen, nitrogen and sulfur, the actual values agreed closely with the theoretical values.

EXAMPLE 2 Preparation of 2-( Z-Methylamino-S-thiadiazolyl l -methyl-5- nitroimidazole A suspension of 20 gm. of ferric ammonium sulfate dodecahydrate in 100 ml. of water is warmed on a steam bath until the solid is completely dissolved. To the above solution 9.6 gm. 0.039 mole, of l-methyl-S- nitro-2-imidazolecarboxaldehyde-4 -methyl-3 -thiosemicarbazone is added, and stirred on the steam bath for an additional 90 minutes. A warm solution containing 59 gm., 0.122 mole of ferric ammonium sulfate dodecahydrate in 300 ml. of Water is added to the above mixture. The mixture is stirred on the steam bath for an additional 2 hours and filtered. The precipitate is washed with water and acetone to give 6.9 gm. ofa yel- 6 low solid having a melting point of from 230 to 23 8C. Recrystallization from 700 ml. of hot ethyl acetate gives 3.3 gm. of bright yellow needles having a melting point of 238C.

EXAMPLE 3 Preparation of 2-( 2-Dimethylamino-5-thiadiazolyl l -methyl-5- nitroimidazole A mixture of 14.1 gm., 0.1 mole, of 1,2-dimethyl-5- nitroimidazole, 12 gm., 0.1 mole, of selenium dioxide and ml. of diethyleneglycol dimethyl ether is refluxed while stirring magnetically for approximately 4 hours. The mixture is then cooled and filtered through a layer of diatomaceous earth into a warm solution of 1 1.5 gm., 0.1 mole of 4',4'-dimethyl-3'-thiosemicarbazide in 50 ml. of water and 10 ml. of glacial acetic acid. A precipitate forms immediately. The mixture is cooled and filtered to give 7.5 gm. of yellow crystals, which have a melting point of from 205 to 210C. The crystals are recrystallized twice from methyl cellosolve and once from ethanol, to give 1.3 gm. of orange needles having a melting point of from 208 to 210C. (decomposition).

A warm suspension of 9.2 gm. of l-methyl-5-nitro-2- imidazolecarboxaldehyde 4 ,4'-dimethyl-3 -thiosemicarbazone (prepared above) in 100 ml. of water is added to a warm solution of 69.4 gm. of ferric ammonium sulfate dodecahydrate in 400 ml. of water. The mixture is magnetically stirred on a steam bath for approximately 4 hours, and filtered. The precipitate 'is washed with water and acetone giving 8.6 gm. of a brown solid having a melting point of from 238 to 240C. (decomposition). Two recrystallizations from N,N-dimethylformamide gives 3.6 gm. of yellow crystals having a melting point of 252C.

EXAMPLE 4 Preparation of 2-( 2-Ethylamino-S-thiadiazolyl 1 -methyl-5- nitromidazole and 2-(2-Dimethylamino-S-thiadiazolyl)-1-ethyl-5- nitroimidazole EXAMPLE 5 Preparation of 2-( 2-Formamido-S-thiadiazolyl l -methyl-5- nitroimidazole The above compound is prepared by refluxing 8 gm. of 2-( 2-amino-5-thiadiazolyl l -methyl-5- nitroimidazole in 30 ml. of 98% formic. acid for 10 hours, cooling, and adding the mixture to saturated so- 7 dium bicarbonate solution, and the solid collected. The compound melts at 225 to 227C.

EXAMPLE 6 Preparation of 2-( 2-Acetamido-5thiadiazolyl l -methy1-5- nitroimidazolc The above compound is prepared by heating under reflux for 30 minutes a mixture of 14 gm. of 2-(2- amino-5 -thiadiazolyl l -methyl-5-nitroimidazole in 280 ml. of acetic anhydride. The mixture is evaporated to-dryness, and the solid residue is washed thoroughly with ether giving 16.4 gm. of a yellow solid having a' m'elting'point of 235C. (decomposition.

EXAMPLE 7 Preparation of l-( 2-Acetoxyethyl )-5-nitro-2-imidazolecarboxalde hyde A 6.27 gm. portion of l-(2-acetoxyethyl)2-hydrox ymethyl-S-nitromidazole is refluxed with 13.3 gm. of

lead tetraacetate in 200 ml. of benzene for 18 hours,-

" hyde.

EXAMPLE 8 Preparation of 2-( 2-Amino-5-thiadizolyl l -(.2-hydroxyethyl )-5- nitroimidazole A 14.25 g. sample of the aldehyde prepared in Example 7 is treated with 5.72 of thiosemicarbazide in 150 ml. of 95% ethanol containing a drop of concentrated hydrochloric acid and the mixture is heated on a steam bath for 20 minutes. The hot solution is filtered to remove insoluble materials, cooled and the yellow-brown crystals are collected. The yield of l-(2-acetoxyethyl)- 5-nitro-2-imidazolecarboxaldehyde thiosemicarbazone is 18.8 g. after drying in a vacuum oven at 60 for 2 /2 hours. Recrystallization of the product gives a yellow solid, melting point 18 ll83.5C.

The thiosemicarbazone (l2.g) is added to 77 g. of ferric ammonium sulfate dodecahydrate in 500 ml. of water at 60 and the mixture is heated to 90l00 for 4 hours. The mixture is cooled, the solid collected and washed with water. Only 0.92 g. (melting point 24925 1) of product is soluble in a large volume of acetone. The remaining product is dissolved in 150 ml. of dimethylformamide, filtered and the filtrate evaporated to dryness to give a solid. This solid is treated with about 20 ml. of acetone, slurried, cooled and collected to give yellow crystals. After drying in an air stream overnight, 55 g. (melting point 253.5255) of 2-(2- amino-5-thiadiazolyl l 2-hydroxyethyl )-5- nitroimidazole is obtained; no carbonyl absorption band is present in the infrared spectrum. Thus, it is not necessary to perform a separate hydrolysis step for the removal of the acetyl group.

EXAMPLE 9 Preparation of 2( 2-Dichloroacetamido-5-thiadiazolyl l -methyl-5- nitroimidazole EXAMPLE 10 Preparation of r 2 2-Amino-5-oxadiazolyl l -methyl-5-nitroimidazole Method A In 180 ml. of hot water, 9.3 grams (0.06 mole) of lmethyl-5 nitro-2-imidazolecarboxaldehyde is slurried while 6.7 grams (0.06 mole) of semicarbazide hydro chloride is added portionwise. After 15 minutes of heating, the mixture is cooled in a refrigerator overnight.-The solid is then collected and washed with water and methanol, respectively, to give a yellow product, melting point 272273C (dec. After drying at 100 for 2 hours under reduced pressure, 11.94 grams of l-methyl-5-nitro-2-imidazolecarboxaldehyde se'rriica rbazone is obtained. This semicarbazone (6.35 grams or 0.03 mole) is added to 10 grams of anhydrous sodium acetate in 50 ml. of glacial acetic acid and 1.25 ml; of bromine is added with continuous stirring. The mixture is heated gradually to give a nearly clear red solution at 50C. which becomes increasingly turbid with time. After heating at for 3 hours, the mixture is cooled and poured on ice. The ,yellow solid is collected, washed with water, then with methanol, and dried under reduced pressure at 70C. for 3 hours to give 5 grams of crude 2-amino-5-( l-methyl-Snitro-Z- imidazolyl)oxadiazole. melting point- 284-287C. (dec.). This material is dissolved in boiling dimethylformamide, ethanol added, and the mixture cooled to give yellow crystals, melting point 29l293C. (dec.). METHOD B I In 25 ml. of methanol, 0.93 grams (5 mmole) of lmethyl-5-nitro-2-imidazolecarboxylic acid hydrazide and 0.53 grams (5mmole) of cyanogen bromide are refluxed for 2 hours, cooled, and poured on ice to give a pale yellow solid. This solid is collected, washed with water, and dried under reduced pressure at C. for 2 hours to give 0.65 grams, melting point 286288C. (dec. of 2-amino-5-( l-methyl-5-nitro-2-imidazolyl- )oxadiazole.

' EXAMPLE ll Preparation of 2-( 2-Methylamino-5-oxadiazolyl l -methyl-5- nitroimidazole To a solution of 8.5 grams (0.055 mole) of-l-methyl- 5-nitro2-imidazolecarboxaldehyde in 50 ml. of ethanol is added a solution of 5.0 grams (0.056 mole) of 4- methylsernicarbazide in 25 ml. of ethanol and 10 ml. of water containing 2-4 drops of concentrated hydrochloric acid. The resulting solution is heated at 60-70C. until a yellow solid starts to separate and then stored at C. for 1 hour; 12.2 grams (98%) of l-methyl-S-nitro- 2-imidazolecarboxaldehyde-4-methylsemicarbazone is obtained melting at 221223C.

Seven grams (0.031 mole) of 1-methyl-5nitro-2- imidazolecarboxaldehyde-4-methylsemicarbazone is dissolved in 85 ml. of glacial acetic acid, 16 grams of anhydrous sodium acetate added, and a solution of 2.5 ml. of bromine in 20 ml. of glacial acetic acid added. The resulting mixture becomes a clear solution as it is heated at 7080C. for 2-4 hours. The solvent is evaporated under reduced pressure and the residue treated with shaved ice until a yellow suspension is obtained. The product is collected, washed with cold water, methanol, and finally with ether. Recrystallization from ethanol containing N,N-dimethylformamide affords 2.7 grams (39%) of a yellow product melting at 23924lC. In another preparation, 63% yield of product melting at 237239C. is obtained without recrystallization.

EXAMPLE 12 Preparation of 2-( 2-Dimethylamino5-oxadiazolyl l -methyl-5- nitroimidazole A solution of 780 mg. (5 millimoles) of Lmethyl-S- nitro-Z-imidazolecarboxaldehyde in ml. 95 of ethanol is treated with 520 mg. (5-millimoles) of 4,4-dime thylsemicarbazide and one drop of concentrated hydrochloric acid to give a yellow solid. The mixture is heated at 40C. for 5 minutes, cooled to 0C., and l 180 mg. (98%) of crystalline l-methyl-5-nitro-2- imidazolecarboxaldehyde 4,4dimethylsemicarbazone is obtained. Recrystallization from ethanol containing N,N-dimethylformamide affords 900 mg. (74%) of material melting at 206-208C.

The procedure is Method A of Example 10 is followed except that 7.5 grams (0.31 mole) of l-methyl-5- nitro-2-imidazolecarboxaldehyde 4,4'dimethylsemicarbazone, prepared above, is used. The product is recrystallized from ethanol to give 5.2 grams (69%) of material melting at 180182C.

EXAMPLE 13 Preparation of 2-( 2-Acetamido-5-oxadiazolyl 1 -methyl-5- nitroimidazole Four grams of acetyl chloride are slowly added to 3.6 grams (0.017 mole) of 2-(2-amino-5-oxadiazolyl)-lmethyl-S-nitroimidazole suspended in a mixture of 44 ml. of pyridine and 35 ml. of benzene. The addition is completed in 30 minutes, the mixture heated at 60-70C. for 10 minutes, and then poured into 500 ml. of ice and water. After the resulting mixture is stirred for 1 1/2 hours, the product separates. Recrystallization from 150 ml. of acetone containing some N,N- dimethylformamide affords 2.4 grams (56%) of pale yellow product. A second recrystallization from acetone gives 2.1 g. of product, melting point 224225C.

EXAMPLE 14 Preparation of 2 2-[ Dimethylaminomethylene) amino -5-thiadiazolyl} 1 -methyl-5nitroimidazole Phosgene gas is bubbled into 100 ml. of N,N-dimethylformamide at 5-l0C. until 2.0 g. (0.02 mole) is absorbed and a crystalline suspension is formed. This suspension is added in portions to a stirred mixture of 4.5

g. (0.02 mole) of 2-(2-amino-5-thiadiazolyl)-l-methyl- S-nitroimidazole and ml. of N,N-dimethylformamide at 25C. After 30 minutes the reaction mixture is diluted with 200 ml. of diethyl ether, and the pale yellow solid is collected, washed with ether, and dried. Treatment of this material with 150 m1. of water and drying affords 5.0 g. (89%) of brilliant yellow solid melting at 230-232C.

EXAMPLE 15 Preparation of 2-[2-( 4-Carbethoxyl -piperazinyl )-5-thiadiazolyl]- l methyl-S-nitroimidazole The compound 1-methyl-5-nitro-2-imidazole carboxaldehyde-4-carbethoxyl -piperazine thiocarbohydrazone is prepared by the procedure of Example 3, 23.2 gm., 0.1 mole, of 4-carbethoxypiperazine-lthiocarbohydrazide replacing the 4,4-dimethyl-3-thiosemicarbazide. The yield is 8.6 gm. and the melting point 182-183C.

A suspension of 6.4 gm., 0.0174 mole, of l-methyl-S- nitro-Z-imidazolecarboxaldehyde-4-carbethoxy- 1 -piperazine thiocarbohydrazone (prepared above) in 200 ml. of boiling ethanol is stirred as a solution of 35 mg. of ferric ammonium sulfate dodecahydrate in 200 ml. of hot water is added in one portion. A deep red-brown solution results. After stirring and heating on the steam 7 bath for four hours, a precipitate is present. This is collected, washed with water, dried and recrystallized from hot ethanol to give 3.6 grams of the title compound melting at l7918lC.

EXAMPLE 16 Preparation of 2-[2-( Hydroxyethylamino )-5-thiadiazolyl l -methyl-5- nitroimidazolc A solution of 1.3 gm., 0.0058 mole, of 2-(2-amino-5- thiadiazolyl)-1-methyl-5-nitroimidazole is dissolved in 25 ml. of concentrated hydrochloric acid, cooled to 5C., stirred, and treated during 5 minutes with a solution of 0.5 gm., 0.0073 mole, of sodium nitrite in 2 ml. of water. The mixture is kept at room temperature for 18 hours. The precipitate present is collected, washed with water, dried and then extracted with warm acetone. Removal of the acetone leaves a solid residue which is recrystallized from a mixture of acetone and diethylether to give 0.16 gm. of 2-(2-chloro-5- thiadiazolyl)-l-methyl-5-nitroimidazole as yellow crystals, melting at 137C.

A mixture consisting of 4.9 gm., 0.02 mole, of 2-( 2- chloro-S-thiadiazolyl l -methyl-5-nitroimidazole 3 .0 gm., 0.05 mole, of ethanolamine, and 50 ml. of p-dioxane is stirred at room temperature for 24 hours. The precipitate is collected, washed with aqueous sodium bicarbonate solution, dried and recrystallized from methanol to give the pure compound melting at 208-209C. Working up the mother liquors gives additional material, the total yield being 3.4 gm.

EXAMPLE 1 7 Preparation of 2- 2-( 3-Dimethylaminopropylamino )-5-thiadiazolyl l -methyl-5-nitroimidazole A mixture consisting of 3.8 gm., 0.0155 mole of 2-(2- chloro5-thiadiazolyl l -methyl-5-nitroimidazole, prepared as in Example 16 2.5 gm., 0.029 mole of 3-dime- ,thylaminopropylarnine, 2.1 gm. 0.025 mole, of sodium bicarbonate and 100 ml. of benzene is refluxed for 22 hours. The reaction mixture is cooled to room temperature, washed with aqueous sodium bicarbonate and sodium chloride solutions. Cooling to C. gives a crystalline precipitate which is collected and recrystallized from benzene to give the pure compound melting at l53l54C. More product is obtained from the mother liquors to give an overall yield of 3.0 gm.

EXAMPLE 1 8 Preparation of 2-( 2-Amino-5-oxadiazolyl l 2-hydroxyethyl )-5 nitroimidazole, 2-( 2-methylamino-5 -oxadiazolyl l-( 2-hydroxyethyl 5 -nitroimidazole and 2-( 2-dirnethylamino-5 -oxadiazolyl l 2-hydroxyethyl)-5-nitroimidazole l-( 2-hydroxy )-5-nitro-2-imidazolecarboxaldehyde l mole) is treated with 1 mole of semicarbazide hydrochloride in ethanol to give nearly a quantitative yield of semicarbazone. The product is treated with bromine and sodium acetate in the manner described in Method A of Example 1 to give the first compound above which meltswith decomposition at 228230C.

The use of 4-methylsemicarbazide and 4,4-dimethylsemicarbazide hydrochlorides instead of semicarbazide hydrochloride, followed by treatment of the methylated semicarbazones with bromine and sodium acetate as above, gives, respectively, 2-(2-methylamino-5- oxadiazolyl l-( 2-hydroxyethyl )-5-nitroimidazole, melting at 202.5204C., and 2-(2'dimethylamino-5- oxadiazolyl )-l 2-hydroxyethyl )-5-nitroimidazole, melting at l7ll73C.

EXAMPLE 19 Preparation of 2-( 2-Amino-5-thiadiazolyl l 2-acetoxyethyl )-5- nitroimidazole The sample (0.1 g.) of 2-amino-5-[l-(2-hydroxyethyl)-5-nitro-2-imidazolyl]thiadiaz0le is dissolved in 2 ml. of hot glacial acetic acid and a drop of concentrated sulfuric acidis added. The solution is refluxed for 45 minutes, cooled and poured on ice to give a yellow solid. This solid is collected, washed with water and dried; the yield is 0.1 g., melting point l59l62C. (turbid). A purified sample of the 2-amino-5-[l-(2- acetoxyethyl)-5-nitro-2-imidazolyl]thiadiazole, melts at l64l65.5C.

EXAMPLE 20 Preparation of 2-( 2-Acetamido-S-thiadiazolyl l-( 2-acetoxyethyl )-5- nitroimidazole The same (0.1 g.) of 2-amino-5[ l-(2-hydroxyethyl)- 5-nitro-2-imidazolyl]thiadiazole is added to 1.5 ml. of acetic anhydride and heated under reflux for 20 minutes. After cooling, the mixture is evaporated to dryness to give a tan solid which is slurried with ether and collected, melting point 258265C.; the yeild is 0.1 l g. This solid is recrystallized from acetone to give the purified product, melting point 264268C.

EXAMPLE 2;

Preparation of 2-( 2-Amino-5-thiadiazolyl l methyl-5-nitroimidazole Hydrochloride Four grams of 2-(2amino-5-thiadiazolyl) l -methyl-5- nitroimidazole is added to concentrated hydrochloric acid. The resultant precipitate of the hydrochloride is filtered off and air dried; melting point 249C. with decomposition. When the hydrochloride is added to water, the free base is formed again without the necessity of using alkali.

EXAMPLE 22 I Preparation of v 2-( 2-Hydroxymethylamino-S-thiadiazolyl)- l -methyl-5- nitroimidazole A suspension of 5 g. of 2-( 2-amino-5-thiadiazolyl)-lmethyl-S-nitroimidazole in 200 ml. of 36% aqueous formaldehyde solution is stirred for 20 hours at room temperature. The suspension is cooled in ice, and the insoluble solid is collected by filtration and washed with acetone. The yield of the title compound is 6.15 g., melting point 177c. with decomposition (rapid heating).

EXAMPLE 23 Preparation of 2-( 2-Amino-5-thiadiazolyl l -ethyl-5-nitroimidazole A slurry of B-( l-ethyl'-5-nitro-2-imidazolyl)styrene (42.7 g. or 0.175 mole) in 350 ml. of methanol containing 14.6 ml. of water at 25 C. is treated with ozone until a nearly clear, pale-yellow solution is obtained. Subsequently, the mixture is treated with 42 g. of sodium iodide in 138 ml. of water and 20.3 ml. of glacial acetic acid at 25C. The mixture is stirred for 40 minutes and 44.1 g. (0.288 mole) ofsodium thiosulfate in 242 ml. of water added. The mixture is filtered, and the filtrate concentrated at 75C. under 15-20 mm. of pressure to give 450 ml. of solution. The solution is acidified with 50 ml. of 6N hydrochloric acid and the benzaldehyde removed at 7075C. under 15-20 mm. of pressure. The residue is then neutralized with saturated sodium bicarbonate solution and extracted with ethyl acetate to give an 81.5% yield of solid aldehyde, melting point 6l67C. after stripping. A sublimed sample of the aldehyde melts at 68-69C. with softening at 65C. When the aqueous ozonized solution, after benzaldehyde is removed, is treated with thiosemicarbazide and mineral acid or semicarbazide hydrochloride and heated, the l-ethyl-5-nitro-Z-imidazolecarboxaldehyde thiosemicarbazone, melting point 241C., or semicarbazone, melting point 223-226C., is obtained. The compound B-( l-ethyl-5-nitro-2-imidazolyl)styrene is prepared by treating l-ethyl-2-methyl-5-nitroimidazole with benzaldehyde in absolute ethanol and potassium tertiary butoxide in nitrogen atmosphere below 37C. A sample purified from 95% ethanol melts at l36.5-137.5C.

A solution of 184 g. of ferric ammonium sulfate dodecahydrate in 680 ml. of water is heated to 60C. and 23.1 g. of l-ethyl5-nitro-2-imidazolecarboxaldehyde thiosemicarbazone (prepared as above) is added with efficient stirring. The temperature is raised to C. and vigorous stirring continued for 4 hours. The reaction mixture is cooled to 4C. and filtered. The solid is washed with warm water and dried under vacuum at 1 C. and then extracted with 800 ml. of hot acetone, followed by three 300-ml. portions of hot acetone. Cooling the acetone gives a solid product, which is filtered off; melting point 231233C.' A second crop with the same melting point is obtained by evaporating the filtrate and recrystallizing the residue from acetone. The combined yield is 17.0 g. (74.5%).

EXAMPLE 24 Preparation of 2-( 2-Amino-5 -thiadiazolyl l-benzyl-S-nitrdimidazole 0.5 g. (1.64 millimole) of B-( l-benzyl-5-nitro-2- imidazolyl)styrene in 95% aqueous methanol is treated with ozone until a clear solution is obtained. To this solution at C., 0.312 g. (1.64 millimole) of sodium meta-bisulfite (Na S O in 3 m1. of water is added. The mixture is then evaporated to dryness under reduced pressure at 75C. and the solid extracted with ethyl acetate. The ethyl acetate extracts are dried, evaporated to dryness to give a yellow-orange residue (probably bisulfite addition product) which solidifies upon standing. This material is dissolved in ml. of aqueous methanol and 0.15 g. of thiosemicarbazide and a drop of 6N hydrochloric are added. After refluxing for 15 minutes and cooling, 0.25 g., melting point 1931 96C., of 1-benzyl-5-nitro-2-imidazolecarboxaldehyde thiosemicarbazone is isolated. The use of a half of an equimolar amount of sodium meta-bisulfite affords the title aldehyde instead of the bisulfite addition product. When the ethyl acetate extract is evaporated to dryness and dissolved in 75% aqueous ethanol and semicarbazone is added, and the mixture is heated on a steam bath for 10 minutes, and sodium acetate is added, the semicarbazone derivative, melting point 226228C. is obtained. Twelve grams of l-benzyl-5 nitro-Z-imidazolecarboxaldehyde thiosemicarbazone are added to a stirred solution of 76.2 g. of ferric ammonium sulfate dodecahydrate in 600 mlfof water at 50C. and the slurry is heated at 90-95C. for 6 hours. The mixture is cooled in ice and the yellow-brown solid collected, washed with water, and dried under reduced .pressure. The yield of product is 10.9 g. Recrystallization from methanol gives analytically EXAMPLE Preparation of I 2-( 2-Amino5-oxadiazolyl)- l -ethyl-5-nitroimida1ole A slurry of 10.2 g. of 1-ethyl-5-nitro-2-imidazolecarboxaldehyde semicarbazone (prepared as in Example 23), and 15.0 g. of anhydrous sodium acetate in 75 ml. of glacial acetic acid is treated with 1.87 ml. of bromine at 81C. under agitation. Stirring is continued at 7580C. for 17% hours, at which time workup of a sample shows starting material still present. An additional 7.5 g. of sodium acetate and 1.00 ml. of bromine are added and stirring continued for 3 hours. A final 3.85 g. of sodium acetate and 0.50 ml. of bromine are added and the reaction mixture is stirred for one hour at 75C. The reaction mixture is cooled to 15C. and poured over ice. The product is filtered off, washed with water and dried under reduced pressure. The yield is 6.3 g. and the melting point 265C. with decomposition.

EXAMPLE 26 ture is heated at 6570C. for 4 hours, then poured on ice and the yellow solid collected, washed with water, and dried under reduced pressure. The yield is 5.7 g. Recrystallization from acetone gives analytically pure material melting at 261.5262.5C.

EXAMPLE 27 Preparation of '2-[ 2-( 2,2Diethoxyethylamino -5 -thiadiazolyl]-1- methyl-S-nitroimidazole A mixture composed of 9.8 g. of 2-(2-chloro-5- thiadiazolyl)-1-methyl-5-nitroimidazole (Example 16), 1 1.0 g. of 2,2-diethoxyethylamine, and 250 ml. of'

dioxane is stirred and heated on the steam bath for 12 hours. It is then diluted with 500 ml. of cold water, cooled, and the precipitated yellow solid collected and dried. Recrystallization from 50% aqueous ethanol gives the pure compound melting at 154155C.'

EXAMPLE 28 Preparation of 2-( 2-Piperidino-5-thiadiazolyl l -methyl-5-' nitroimidazole A mixture composed of 7.4 g. of 2-(2-chloro-5- thiadiazolyl)- l -methyl-5-nitroimidazole (Example 16),

6 g. of piperidine, and 200 ml. of dioxane is stirred at room temperature for 24 hours, and then taken to dryness under reduced pressure. The residue is shaken with m1. of water and the insoluble portion collected and dried. Recrystallization from 150 m1. of 2- methoxyethanol gives 7.6 g. of the subject compound melting at 212213C.

EXAMPLE 29 Preparation of 2-( 2-n-Hexylamino-S-thiadiazolyl 1 -methyl-5 l nitroimidazole The preparation of the title compound is carried out essentially as described for the Z-piperidino derivative (Example 28), 6.5 g. of n-hexylamine replacing the piperidine. The crude product is recrystallized from ml. of 2-methoxyethanol to give 5.2 g. of the pure compound melting at 146147C.

EXAMPLE 30 Preparation of 2-[4-( 2-l-lydroxyethyl l -piperazinyl-5-thiadiazolyl]- l methyl-S-nitroimidazole The preparation of the subject compound is carried out essentially as described for the 2-piperidino derivative (Example 28), 3.9g. of 1-(2-hydroxyethyl)piperazine replacing the piperidine. After recrystallization from ethanol, 3.5 g. of pure compound is obtained, melting at 204206C.

EXAMPLE 31 Preparation of 2- 2-[ 4-( 3-Dimethylaminopropyl l -piperidino -5- thiadiazolyl l -methyl-5-nitroimidazole The preparation of the title compound is carried out by the procedure described for the Z-piperidino derivative (Example 28), 5.1 g. of 4-(3-dimethylaminopropyl)piperidine replacing the piperidine. Recrystallization from methanol gives 5.6 g. of pure product, melting at l74l75C.

EXAMPLE 32 Preparation of 2- {2-[N-( 2-Hydroxyethyl )methylamino]-5-thiadiazolyl l -methyl-5-nitroimidazole The preparation of the above compound is carried out essentially as described for the Z-piperidino derivative (Example 28), an equivalent of N-methylethanolamine replacing the piperidine. The pure compound melts at 158l60C. after recrystallization from methanol.

EXAMPLE 33 Preparation of 2-[2-( l-Piperazinyl )-5-thiadiazolyl]- l -methyl-5- nitroimidazole The preparation of the subject compound is carried out essentially as described for the 2-piperidino derivative (Example 28), two equivalents of piperazine replacing the piperidine. The crude product is recrystallized from Z-methoxyethanol to yield the pure compound melting at 24024lC.

EXAMPLE 34 Preparation of 2-[2-( 4-Methyll-piperazinyl )-5-thiadiazolyl 1 -methyl-5-nitroimidazole The preparation of the above compound is carried out in the manner described for the 2-piperidino derivative (Example 28), an equivalent of l-methylpiperazine replacing the piperidine. The crude product is purified by recrystallization from Z-methoxyethanol and melts at 242244C.

EXAMPLE 35 Preparation of 2-{ 2-[4-( 3-Dimethylaminopropyl l -piperazinyl]-5- thiadiazolyl l -methyl-5-nitroimidazole EXAMPLE 36 Preparation of 2-{2-[4-(2-Pyridyl)- l-piperaziny1]-5-thiadiazolyl}- l methyl-S-nitromidazole The preparation of the above compound is carried out essentially as described for the 2-[4-(3-dimethylaminopropyl)l-piperazinyl derivative] (Example 35), an equivalent of l-( 2-pyridyl)piperazine replacing the 1-(3-dimethylaminopropyl)piperazine. After recrystallization from 2-methoxyethanol, the pure compound melts at 280-282C.

EXAMPLE 37 Preparation of 2-{2-[4-(2-Thiazolyl)-1-piperazinyl]-5-thiadiazolyl}-lmethyl-S-nitroimidazole The preparation of the subject compound is carried out essentially as described for the 2-[4-(3-dimethylaminopropyl)-l-piperazinyl] derivative (Example 35), an equivalent of l-(2-thiazolyl)piperazine replacing the l-(3-dimethylaminopropyl)piperazine. After recrystallization from Z-methoxyethanol, the pure compound melts at 297300C.

EXAMPLE 38 Preparation of 2-( 2-Cyclohexylamino-5-thiadiazolyl l -methyl-5- nitroimidazole A mixture of 5.0 g. of 2-(2-chloro-5-thiadiazolyl)-l- .methyl-5-nitroimidazole, 6.0 g. of cyclohexylamine and v 125 ml. of dioxane is stirred at reflux until thin-layer chromatographic analysis indicates that the chloro intermediate is completely utilized. The dioxane is removed under reduced pressure and the residue triturated with aqueous sodium bicarbonate solution. Recrystallization from a mixture of ethylacetate and acetone yields the pure compound melting at 215217C.

EXAMPLE 39 Preparation of 2-( 2-t-Butylamino-S-thiadiazolyl)- 1 -methyl-5- nitroimidazole The preparation of the above compound is accomplished essentially by the procedure described for the 2-cyclohexylamino derivative (Example 38), an equivalent of t-butylamine replacing the cyclohexylamine. Recrystallization of the crude product from a mixture of ethyl acetate and acetone gives the pure compound melting at 249-25 1C.

EXAMPLE 40 Preparation of 2-( 2-n-Octylamino-5-thiadiazolyl)- 1 -methyl-5- nitroimidazole The preparation of the above compound is carried out essentially as described for the 2-cyclohexylamino derivative (Example 38), an equivalent of n-octylamine replacing the cyclohexylamine. The crude product is purified by recrystallization from a aqueous 2-methoxyethanol and then melts at l30l32C.

EXAMPLE 41 I Preparation of 2-( 2-Chloroacetamido-S-thiadiazolyl l -methyl-- nitroimidazole Twenty grams of 2-(2-amino-5thiadiazolyl)-l-methyl-5-nitroimidazole is added in portions to a solution of 60 g. of chloroacetic anhydride in 250 ml. of dioxane at 90 95C. The mixture is stirred at this temperature for three hours. After standing at room temperature, the mixture is filtered and precipitate washed with diethyl ether and dried. Recrystallization from 2-methoxyethanol gives the pure compound, melting at. 239240C, with decomposition.

EXAMPLE 42 Preparation of 2-( 2-Morpholinoacetamido-5 -thiadiazolyl 1 -methyl- 5-nitroimidazole EXAMPLE 43 Preparation of 2-( 2-Amino-5-thiazolyl l -propyl-5 -nitroimidazole The procedure of Example 23, with the substitution of /3-(l-propyl-S-nitro-2-imidazolyl)styrene for B-(lethyl-5-nitro-2-imidazolyl)styrene in the first step is utilized to prepare 2-(2 amino-5-thiadiazolyl)-l-propyl-S-nitroimidazole, melting at 234.5236C.

EXAMPLE 44 Preparation of Chick Diet The following feed composition is employed in all the the poultry experiments hereinafter set forth except where otherwise stated:

Vitamin Pre-Mix 0.5 Trace Minerals 0.1 Sodium Chloride 0.3 Dicalcium Phosphate 1.2 Ground Limestone 0.5 7:- Stabilized Fat 4 Dehydrated Alfalfa. I77: 7 2 Corn Gluten Meal, 41% 5 7r Menhaden Fish Meal, 60% 5 70 Soybean Oil Meal, 44% 30 7c Ground Yellow Corn, To I00 Fine The vitamin pre-mix in the above feed composition is prepared from the following formulation. The expressions of quantity relate to units per kilogram of the feed composition.

Butylated Hydroxy Toluene I25 mg. dl-Methione 500 mg. Vitamin A 3300 l.U. Vitamin D '1 100 LU. Riboflavin 4.4 mg. Vitamin E 2.2 LU.

-continued Niacin 27.5 mg. Pantothenic Acid 8.8 mg. Choline Chloride 500 mg. Folic Acid 1.43 mg. Menadione Sodium 1.1 mg. Bisulfate Vitamin B ll mcg. Ground Yellow Corn, to 5 gm. Finc EXAMPLE 45 Utilization of compounds of the present invention in controlling colibacillosis This example demonstrates the effectiveness of 2-( 2- amino-5-thiadiazolyl)-l-methyl-5-nitroimidazole, 2-(2- acetamido-S-thiadiazOlyl l -methyl-5-n itroimidazole and other imidazoles in controlling colibacillosis in poultry. y

Three groups of 10 five-day old sexlinked pullet chicks are infected parenterally, in the left thoracic air sac, with 0.2 ml. of a 10 dilution of a Trypticase Soy Broth culture of Escherichia coli, the causative agent-of colibacillosis in poultry. The compounds to be tested are administered by gavage as a single oral dose in an aqueous solution or suspension, and the chicks are permitted to feed ad libitum the feed composition prepared in Example 44. Twelve days after treatment, the test is terminated and the number of survivors in each group recorded. The results are compared with two control groups of 20 chicks each, in which one control group is infected and untreated, and the second control group is uninfected and untreated. The results of the test are set forth in the following table:

Dose is in terms of milligrams per kilogram of body weight.

Four groups of 40, five-day old sex-linked pullet chicks are infected in the same manner as herein set forth with Escherichia coli. The compound 2-(2- acetamido-5-thiadiazolyl)-1-methyl-5nitroimidazole is administered by gavage as a single oral dose in an aqueous medium, and the chicks are permitted to feed ad libitum the feed composition prepared in Example 44.

Twelve days after treatment, the test is terminated and the number of survivors in each group recorded. The results are compared with two control groups of 40 chicks each, in which one control group is infected and untreated, and the second control group is uninfected and untreated. The results of the test are set forth in the following table.

TABLE ll TABLE Ill-continued Compound Dose* Total Chicks Survivors Compound Dose* Total Chicks Survivors T d Tested Z-(Z-acetamido- 40 mg. 40 39 5 0.006% l 2 S-thiadiazolyl 2 2-[N-( Z-Hydroxy- 0.1 10 8 l:me thyl-- 20 mg. 4() 35 ethyl )methy]amino] n1tro1m1dazole 5-thiadiazolyl}- l l0 mg. 40 29 methyl-S-nitrd- Control imidazole 2-( 2-Morpholino- 0.1 7: l0 l0 Infected Untreated 40 3 acetamido-5-thiadia- Uninfected Untreated 40 40 1 1 1 5. 0 025% 5 1 nitroimidazole Dose is 1n terms of milligrams per k1logmm of body e1ght. L0 l0 l0 diazolyl 1 -methyl- 5-nitroimidazole 0.025% 5 3 hydrochloride M EXA PLE 46 2-[2-(4-Methyl-l- 0.1 10 4 Utilization of compounds of the present invention in ggg 'f controlling fowl typhoid methyl-S-Kitroimidazole This example demonstrates the effectweness of the 2 (2 Ethylamino 5 0'1 w 10 compounds of this 1nvent1on. thiadiazolyl)-l- Groups of five one-day old sex-linked pullet chicks figgj 002567 10 3 are infected orally by gavage with 0.5 ml. of a 10 dilu- 5 3 tion of a five-hour Trypticase Soy Broth culture of Sal- Z-( Z-CyClOlICX yl- 0.1 10 x monella gallinarum, the causative agent of fowl tyzolyl 1 -methylphoid. Each chick receives approximately 6 X 10 via- 5.nitroimidaz0|e ble cells. Medication is administered continuously in 2-(2-t-Butylamino- 0.1% 10 6 the feed, beginning 3 hours before infection and conq T methyl-5-mtro1m1- tinumg for 10 days, at which time the test is termlnated m and the number of survivors in each group recorded. 2-(2-n-Hexylamino- 0.1 10 6 Each test utilizes two control groups of chicks, the first i group comprising 20 chicks which are infected and unimidazole treated, and the second group comprising 10 chicks 2-t2-liperidino-5- 0.1 10 4 which are uninfected and untreated. The results of the tests are set forth in the following Table III (in some iniinidazole stances, the results from two or more tests are com- 2-[2-(l-Piperazinyl)- 0.1 10 8 S-Ihiadiazolyl 1- lblned) methyl-S-nitroimida- 0.025% 10 1 TABLE 111 2(2-am1no-5-th|adi- 0.1 72 l0 10 Compound Dose* Total Chicks Survivors azoly Z- y rOXy- T d ethyU-S-nitroimi- 0.025% 10 8 dazole -A l0 9 2-l4(2-Hydroxy- 0.1 10 7 y l ethyl 1 -piperazinylmtrolmldazolc 0.05 Zr 5 5 S-thiadiazolyll-l- 0.025% l0 3 0025 5 5 methyl-S-nitroimida- 0.006% 5 1 zole 5 5 2-(2-Dichloroace- 0.1 10 7 yU- tamido-S-thiadiw y 5 4 zolyl 1 -methyl-5- 0.025% 15 3 nitroimidazole nitroimidazole l 5 3 2-{2-[(Dimethyl- 0.1 5 5 2-(2 -M ethylam1no- 0.1 5 3 aminomethylene)ami- 5-thiadlazolyl)- no] 5-thiadiazolyl}- 0.05 70 5 5 l-methyl-5 5O l-methyl-S-nitroiminitroimidazole dazole 2-( 2-Dimethyl- 0 1 5 4 2-(2-Formamido-5- 0.1 10 5 arn1no-5-th1athiadiazolyl l diazolyl l methyl-S-nitroimida- 0.025% 15 l l methyl-5-nitro zole imidazole 0.006% 15 9 l0 l0 2-(2-Amino-5-oxadia- 0.1 10 10 diazolyl l -ethylzolyl I -methyl-5- S-nitroimidazole 0.025% 10 9 nitroimidazole 0.025% l0 l0 2-(2-Amino-5-oxa- 0.1 10 10 0.006% 10 9 diazolyl)-l-ethyl Z-(Z-Dimethylamino- 0.1 7c 10 9 S-nitroimidzizole 0.025% 5 5 -5-0 adiaz ]yl) 1 0.006% 5 3 methyl-S-nitroimida- 0.025% 10 9 Z-(Z-AminO-S-thia- 0.1 10 10 zole 'diazolyl l -benzyl- 0.006% l0 2 S-nitroimidazole 0.025% 10 4 2-(2-Acetamido-5-oxa- 0.1 7: l0 l0 2- 2-Amino-5-oX=i- 0.1 10 9 fl yp y diazoxyl I 5-n1tro1m1dazole 0.025% 10 8 S-nitroimidazole 0.025% l0 1 2-(2Methylamino-5- 0.1 5 5 0.006% 10 1 oxadiazolyl)-l- 2 (2 Hydroxymethy] OJ 10 10 methyl-S-nitroimida- 0.025% 10 10 amino-S-thiadiazozole lyl 1 -met1i 1-5- 0.025% 10 2 0.006% 10 g nitroimidazole 2-(2-Methylamino-5- 0.1 l0 6 Dose in in terms of percentage by weight of the feed eompmitim prepared In Example 45.

EXAMPLE 4'7 Utilization of composition of the present invention in controlling enteritis This example demonstrates the effectiveness of 2-(2- amino-5-thiadiazolyl)-l-methyl and l-ethyl-5- nitroimidazole in controlling enteritis.

Three groups of 10 female Swiss Webster mice weighing 20 gm. are infected intraperitoneally with 0.5 ml. of i" dilution of a -hour Trypticase Soy Broth culture of Salmonella c/roleraesut's var. kunzendorf, the causative agent of enteritis in pigs, an organism originally recovered from a field outbreak of Salmonella c/toleraesuis var. kunzendorf in pigs. Each mouse receives approximately 4.6 X cells as the inoculating dose.

The mice are fed a medicated feed, which is a commercial mouse chow containing the compounds to be tested for 3 hours before infection until 7 days after infection. The mice are held for an additional 7 days after the medication is stopped, and the number of survivors in each group recorded. The midicated feed is prepared by thoroughly admixing calculated amounts of the compounds with commercial mouse chow to provide essentially uniform distribution in the feed offered. The above results are compared with two control groups of 10 mice each. in which one control group is infected and untreated, and the second control group is uninfectcd and untreated. The results of the test are set forth in the following table:

Dose is in terms of percentage by weight of the commercial mouse chow.

EXAMPLE 48 Experiments were conducted to determine the antibacterial activity of compounds of the present invention against Proteus mirabt'lls. Staphylococcus aureus strain Smith and Staphylococcus aureus strain Rose a tetracycline-resistant strain of Staphylococcus tum-u. Infection was initiated in Carworth Farms CFl-s fe male mice (body weight l822 mg.) by intra-abdominu infection of 0.5 ml. of the broth dilutions of 5 hour cul tures of the above organisms. The results Obtained ar summarized in the following Tables V. VI, and VII.

TABLE V Compound Results alive/tested 2-(2-Amino-5-thiadiazolyl I -mcthyl-5- nitroimidazole 2-( Z-Methylamino-S- thiazolyl l -mcthyl- S-nltroimidazole 2- 2-|(Dimethylamlno methylene)amino]-5-thiadiazolyl -l-mcthyl-5- nitroimidazolc 2-( 2-Formamido-5-thiadiamlyl I -methyl-5- nitroimidazole 2-( Z-Amino-S-thiadiazolyI)-I-methyl-$-nitrm imidazole hydrochloride this test is usually at least TABLE VI Tests with SlupIu-Imxrur aureus strain Smith Dose Results Compound mgJkg. alive/tested 2-( Z-Amino-S-thiadin- 64 l (1/20 zolyl l -methyl-5-nitroimidazolc 2-( 2-Mcthylnmino-5- I 28 4/ 5 thiadiazoIyl)-l-methyl- S-nitroimidazole 2-I 2-(4-Carbethoxy- I 256 8/ I 0 piperazinyl )-5-thiadiazolyl I l -mcthyl-5-nitroimidamlc 2-( Z-Dimethylamino-S- 5 12 4/5 thiadiazolyU-I-methyl- S-nltroimidazole Z-{Z-HDimethyIamino 64 5/5 methyle 1e)amino I-S-thiadiazolyl l -methyl-5-nitroimldazo e 2-( 2-Formamldo-5-thladla- 64 5/5 zolyIl-l-methyl-S-nitrm imidnzoie Z-(Z-Amino-S-oxadiazolyl I28 9/10 l-methyl-5- nltroimidazole 2-( 2-Acetamido-5-oxadia- 122i 3/5 zolyI)-l-methyl-5-nitroimidazole 2-( Z-MethylamlnoS-oxadia- I28 3 /S zolyl l -methyl-5-nitroimidazole 2-( 2-Amlno-5-oxadlazolyl l I28 3/ 5 eth'yl-S-nitrolmldazole 2-( Z-Amino-S-thiadiazolyl I28 5/5 l-methyl-S-nltrolmidazole hydrochloride 2- {2-[4-( 3-DimethyIamlno- 256 3/ 5 propyl I -piperazinyl I-S-thiadiazolyl}- l -methyI-5-nitrolmidazole The mortality among infected untreated controls in this'test is usually at least 95' thladlazolyI)-l-methyl- 3,904,756 23 24 TABLE VILcominued The mortality among infected untreated controls in this test IS usually 95% or more. Activity was also displayed by the compounds of this invention against the Tests with Sluphylmrxrux uun'm strain Rose Dose Compound mg./kg Results alive/tested following organisms in mice: E. coli, Salmonella ly- Mhmimidmolc 5 plwsu, Klebslella AD, Aerobacrer aerogenes, and Strep/0 2-(2-[lDimethylamino- 128 10 10 (mus Py mam A methylene)amino]-5-thiadiamlyl}-l-methyl-5- EXAMPLE 49 nitroimiduzole 2 -(2 Formamidofi thia 512 5/5 This example demonstrates the effectiveness of the 10 nitroimidazoles of this invention in controlling a 242-Aminn-5-oxadla- 512 5/5 Pasteurella multocida infection in mice zol l)-l-meth l-S-nitroimiiiuzole y SWISS Webster, female mice weighing 22+2 g. were z-(z-plmcih lmnino-s- 5l2 infected by intrapeutoneal injection of 0.5 ml. of a 5 oxadiazolyU-l-methyl- 1 f smimimidmolc 5 hour Trypttcase Soy Broth culture containing l0 c u. fi t -q f y no-i-oxu- I28 5/5 of P. mulroclda. Each mouse was given by gavage at the zf gz gg sj time of infection 2 mg. of the test drug suspended in 2-(Z-Amino-S-thiadiazolyh- 5:2 5/: 0.2% agar. The number of mice surviving two weeks l"" i"" post-infection over the number of mice tested is re- --(Z-Ammo-S-oxudtazolyh- $l2 5/5 |.clhynsmimimidumk 2o corded below for each compound. The results are shown in Table Vlll.

TABLE Vlli Survlvonfl'otal rug/mouse Compound 2 I 0.5 0.25

2-( 2-Amino-5-thiadiazolyl l0! i O l-ethyl-Smitroimiduzole 2-(2-Amino-$-oxudiuzolyl)- 8/10 1-ethyl-$-nitrnirnidazolc 2-( Z-Arnlno-S-thiadiaznlyl 0/ l (l l-henzyl-S-nitroimidazole 2-(Z-HydroxymethyluminolO/lO S-thiadiuzolyl l -methyI-5- nltroimidazolc 2-{2-[N-(Z-Hydroxyethyhl0/l0 3/10 l/lO methylumlnol-fi-thiadiamlyl? l -methyl-5-nitroimi dam e t 2-( 2-Morpholinoacctamido-fi- 2/l0 thiudiazolyl)-l-methyl-5- nitroimidazolc 2-(2-Ethylamlno 5-thiudiulU/lt) zolyli-l-methyl-S-nitroimldazulc 2-{2-l4-(J-DlmethylaminolU/lt) propyl l -piperazinyi 1-5- thiudiazolyl}-l-mcthyl-S- nitroimiduzole I 2-(Z-n-HcxyluminmS-thiadlat l 9/10 8/l0 2/10 zolyl l -methyl-5-nltroimidumic IZ-(Z-Piperidlno-S-thiadiul v 9H0 4/l0 5/l0 zolyl l -methyl-5-nitroimidumle 2-(2-Amlno-5-oxudluzolyl)-l- IO/l0 9/l0 (2-hydroxyethyl )-$-nltroimldazole 2-{2-l4-(2-Thiazolyh-l-plpera- 2/l0 zlnyl l-S-thiadlazolyl} l -methyl- 5-nltrolmldazole 2-l 2-( l-Plperazlnyl )-5-thlall l0 dlazolyl ll -methyl-5-nitrolmlduzole 2-|4-(2-Hydroxyethyl)-ll0/l0 plperazinyl-S-thladlazolyl l-methyl-S-nitrolmldaznle 2- (2-[ 4-( Ii-DlmethylamlnolO/ l 0 9/ l 0 propyl l -piperidlno ]-S-thladiazolyl}- l -methyl-5-nitroimitluzoie 2-( 2-Amlno-5-thiadiaznlyl l 64/70 5/160 "10/179 87/150 mcthyl-Smitrolmidamlc y 2-{2-[tDimcthylaminomethyl0/l0 28/30 39/40 32/40 lene )nmlno I-S-thiadlazolyl] I -methyl-$-nitroimidazole Z-(Z-FOrmamido-S-thludla- Ill/l0 Hi0 20/20 lh/ZO zolyl l -methyl-5 -nltroimldazole t v r 2-( Amino-S-thiadiamlyU-l- 20/20 Ill/2t) 23/30 (Z-hydmxyethyl )-5-nitmimidazole I Z-(Z-Amlno-Soxudiumlylbl- I W20 3 20 3 20 2/20 nartality among infected untreated controls in this experimental infection is usually at least broth or broth containing Salmonella choleraesuis varil ety kunzendorf organisms. Most of this feed is consumed within an hour. For the rest of the trial the pigs are observed and graded daily. Five hours after infection the pigs are given normal feed or feedcontaining furazolidone, a known antisalmonella agent, or 2-( 2- amino-S-thiadiazolyl l -methyl-5-nitroimidazole.

Medicated feed is employed for 14 days after infection, at which time it is replaced with normal feed. The trial is terminated four weeks after infection. The results of this experiment are shown in Table IX.

TABLE IX Within 1 hour after infection, the mice are treated with drug suspended in 0.5 ml. of 0.2% aqueous agar administered by oral gavage. The drugs are tested at four to six dosage levels with 20 to 30 mice at each level. The results of replicate tests are pooled to determine the median effective dose (ED by the procedure of Litchfield and Wilcoxon, 1949, a simplified method of evaluating dose-effect experiments, J. Pltarmaeol. Exptl. Therap. 96:99-113. Data are tivenin Table X hereinafter.

Concentrations of drug in plasma are determined by assay of blood obtained by heart puncture of noninfected mice. Blood samples (about 0.5 ml, per mouse) from 10 mice receiving the same 'does'ofudrug are pooled. The drug is assayed in the plasma by a standard agar-diffusion plate method with Bacillus subtilis (Lederle No. 17) as the test organism. Peak plasma drug concentrations are given in Table XI hereinafter where they are associated with the median effective dose.

Mortality of Pigs Infected Via Feed with Salmonella ('ltoleraesuis var. kunzendorf and Medicated Via Feed for the First 14 Postinfection Days "During first day postinfection. Differs significantly from infecled untreated group (P .0l

EXAMPLE 5] The compound 2( 2-amino-5-thiadiazolyl)-l-methy1- S-nitroimidazole (Example 1 is active against both gram-negative and gram-positive organisms. In mice, it

is rapidly absorbed, well tolerated and effective against lethal bacterial infections. In vitro, it is effective against gonococci and gram-negative bacilli. Its action is bactericidal.

In the following experiments both the base and the hydrochloride salt of 2-(2-amino-5-thiadiazoly1)-1- 5O methyI-S-nitroimidazole are used. On a molar basis, they are equivalent in activity. In mice, the method used in this experiment has been described. (Redin, G. S., Antibacterial activity in mice of minocyclin, a new tetracycline, Antimicrobial Agents and Chemotherapy- 1966, p. 371-376). Infections are initiated in Carworth Farms CF-l female mice (body weight, 18 to 22 g.) by intra-abdominal injection of 0.5 ml. of the indicated broth dilutions of 5-hour cultures of the following organisms: Klebsiella pneumoniae AD, 10"; Salmonella typhosa 39, unidiluted; Escherichia coli UC3l l, 10*; Aerobacter aerogenes 75, 10 Proteus mirabilis 4671, 10*; Slzz'gella flexneri 2a M22-18, undiluted; Streptococcus pyogenes C203, 10 Staphylococcus aureus 3,

undiluted; S. aureus Rose, undiluted; S. aureus 5, undi 65 luted; S. aureus Smith, 10 and Diploeoceus pneumoniae SVI, 10 For the Neisseria meningitidis N20 infection, 0.5 ml. of a 48-hour broth culture is used.

TABLE x The compound 2-( 2-amino-5-thiadiazolyl)- l -methyl-5-nitroimidazole tested against bacterial infections in mice (single oral dose) Infection" Median effective dose Neisseria meningitidis N20 0 0.2 (0.1-0.5) Klebsiella pneumoniae AD 8 (6-1 I Salmonella typhosa 6539 14 10-19) Escherichia coli 31 l 16 l2-2l Aerobacter aeragenes 54 (43-68) Proteus mirabilis 4671 60 (SO-) Shigella flexneri 2a M22-.I8 89 (52-150) Streptococcus pyogenes C203 4 (36) Stapltvloeocctis aureus 3 14 (9-21 S. aureus Rose 28 18 14) S. aureus 5 46 (26-81 S. aureus Smith 56 (40-77) Diploeoecus pneumoniae SV] 69 (53-90) Mortality rate of the untreated infected control mice was 907: within 3 days. The experimental period was 14 days. "Figures in parentheses indicate 71 confidence limits.

TABLE XI Peak concentrations of test compound in plasma associated with median efiective doses ED Peak concn. Infection (mg/kg.) in plasma (ug/ml.)

Net'sseria meningilidis N20 0.2 1 Streptococcus pyogenes C203 4 Klebsiella pneumoniae AD 8 Salmonella t \'plmsa 6539 14 Escherichia coli 31 l 16 2-4 Staphylococcus aureus 3 I4 Staphylococcus uureus Rose 28 Staphylococcus aureus 5 46 27 TABLE XI-Continued drug that completely inhibited growth is recorded as the minimal inhibitory concentration.

For the comparison of inhibitory and killing concen- Infection (mg/kg.) in plasma (ug/ml. trations, tests are performed in Nutrient Broth. A stock Slaplrvlococcus aureux Smith 56 6-12 SOluPlon of tes t compound In 1 2O Aembueter uerngumkr 75 54 is diluted 1:40 in broth and ster1l1zed by filtration 4 l8 through a O.2p.m membrane filter (Millipore Corp., prom ,nimbilix 4671 .Bedford, Mass). Twofold dilutions of the sterile broth 7 n s m l I) I th H mm, ,d n] 10 solution are inoculated with equal volumes of 10 dilu- ..-21m1no-.- I 1217.0 -mc umr i\'.( L. l

y y tions of a 5-hour broth culture and are incubated at 37C. for 24 hours. Subcultures are made from all clear EXAMPLE 52 I Y tubes by plating 0.1-ml. samples in duplicate and incu- The followmg Orgamsms are used f an P test hating the plates for 24 hours. The following Table XII ing study involving the following species: Neisserza gonsummarizes the results orrhoeue (36 strains), N. meningitidis (24), Shigella flexneri (II); S. sonnei (3), Salmonella typ/zimurium (5 TABLE XII 5. entfrlduls ('1), S mfantls (I), C w erasum Activity of 2-(2-amino-5-thiadiazolyl)-l-methyl-5-nitroimidazole organlenburg l Salmonella species not further identlclinical isolates (agapdilution method) fied (5), E. coli (24), Enterobacter (21 strains of Klebi- No. of Isolates Test Organism Tested Compound" sella pneumonlae and Aerobacter aerogenes), Proteus sp., all indole-negative (20), Pseudomonas sp. (10), D. Gonococcus 36 0.12 pneumoniae 10), Streptococcus pyogenes l5), Staph ygf sgg fi $2 1 lococcus aureus (23), Streptococcus faecalis, enteros u u 14 4 cocci (28). The media used are Nutrient Broth (Difco), 24 8 25 Iznlerobueler 21 8 Nutrient Agar, Nutrient Agar containing 5% defibrlpmum 20 64 nated rabbit blood, or chocolate agarprepared by comf' 10 123 i L GC d- DzpIoeoccus l0 4 bmmg the following materials from BB me rum Swlnmmzm base, hemoglobin, and enrichment supplement. Group A 15 '8 Th ff t' f the a t'bacterial a ents a ainst 23 16 e e ec veness o r1 1 g 3Q Iinlcrococcus 28 64 the clinical isolates is determined by the agar-drlutlon method 2 ino y y "Drug concentration (ug/ml.) required to inhibit 9071 of isolates nitroimidazole is relatively insoluble in water; therefore, a stock solution is prepared in dimethylsulfoxide (DMSO) and serially diluted in DMSO, then 0.1 ml. of EXAMPLE 53 dilution is mixed with 10 ml. of fluid agar. The concen The efficacy of the compounds of the present inventration (1%) of DMSO did not affect the growth of the tion for enhancing the growth rate and for improving test cultures. The plates are inoculated with 10' dilufeed efficiency of animals is shown in the following tions of 5 hour broth cultures by means of a multiple tests. Day-old chicks are divided into groups of 10 birds inocula-replicator. (Steers, E., E. L. Foltz, and B. S. each, five males and five females. These groups are Graves, An inocula replicating apparatus for routine then weighed, placed in pens and given free access to testing of bacterial susceptibility to antibiotics, Anwater and feed. The diet used is the same for both the tibiot. Chemotherapy 91308-31 1 (1959). With menintest groups and the controls except that 100 ppm of 2- gococci and gonococci, 24- to 48-hour surface growth (2-amino-5-thiadiazolyl)-l-( 2-hydroxyethyl)-5- on chocolate agar is Suspended in Trypticase Soy Broth nitroimidazole is admixed with the diet of the test (BBL) for reading of 40 to 50% transmittance at 530 groups. All feed is weighed such that an accurate deternm in a Bausch and Lomb Spectronic-ZO calorimeter; mination as to the amount of gain per unit of feed can the cultures are then diluted 1,000-fold in broth. lnocube acertained. lated plates are incubated at 37 C for 24 hours; menin- All groups are weighed at 0 and 19 days after the test gococci and gonococci are incubated in an atmosphere 50 is begun and the data obtained are reported below. of 5% carbon dioxide. The lowest concentration of Table XIII, along with a description of the diet used.

TABLE XIII Wt. in Grams inc. over No. Days Gain lb, feed/ Controls Supplement Birds 0 19 19 Days lb. gain Gain FIG None 10 43 328.0 285 1.62 None 10 43.1 296.2 253 1.62

None 10 43.1 314.6 272 1.69 None 10 43.1 335.8 293 1.60

Average 43.1 318.7 276 1.63

PPM 2-(2-amino- S-thiadiazolyl)-1- (2-hydroxyethyl) -5-nitroimidazole Average 43.1 340 297 1.525 7.6% 6.5%

DIET

D. Basal CompoundX'(20O gm/ton of feed) E. Basal AUREO S.P25O (100 gm. of chlorotetracycline, 100 gm. of sulfamethazine and 50 gm. of

Ground yellow corn 51,4 soybean on meal (44% 300 penicillin/ton of feed). I Corn gluten meal 5.0 Compound X 2-(2-am1no-5-thlad1azolyl)-l-methyl- Menhaden fist meal (60%) 5.0 idaz Fat (NRC-50) 4.0 5 nmolm Dehydrated alfalfa meal 17% 2.0 The composition of the basal ratlon fed shown in Ground limestone (33% Ca) 0.5 Table XV. The length of these experiments is 42 days. Dicalcium phosphate 1.2 Results. Sodium chloride .3 l0 Trace minerals 0.1 Vitamin remix .5 The average performance data for these two experi- Lime Crest Z42 Delamix which contains 25.57! calcium. 6.07: manganese, 2.071 'ments ls shown In Tabl e galns for iron. 2.0% zinc. 0.12% iodine and 0.02% cobalt three levels of 2-(2-am1no-5-th1ad1azolyl)-l-methyl-S- Provides 3300 LU. Vitamin A. 1100 Lu. Vitamin D1,, 2.2 LU. Vitamin E 500 mg. nitroimidazole clearly Superior to those for the choline chloride, 500 mg., DL-methionine, 125 mg. ethnxyquin. 27.5 mg. niacin. 15' 8.8 mg. pantothenie acid. 4.4 mg. riboflavin. 1.43 mg. folic acid. l.l mg. menadimedlcated control p g The P g g wo mgr Viwmi Bu Par kibgwm feed higher levels 100 and 200 grams per ton) gained more ra idl than those receivin the lower level 50 EXAMPLE 54 p y g gm./ton). Treatment E (AUREO SP. 250) is included In the fOIiOWIHg tests, the same difit and procedure in these experiments as a positive c ntr L were used as described in Example 53 above, excepting I that 10 ppm. of test compound was included in the A marked improvement in feed efficiency was obdiets of test groups. 1 r tained at all levels of 2-(2-amino 5-thiadiazolyl)-1 Data obtained are reported in Table XIV below. methyl-S-nitroimidazole fed.

. TABLE XIV 1 .Weight in Grams 71 Increase Over Days Gain Controls Number i 4 b. d/- Supplement Birds 0 l9 l9 Days 7 lb. gain C iain None l0 40.5 323.5 283.0 1.57 2-(2-Amino-5-thiadiazolyl) 1 1 A l-methyl-S-nitroimidazole 10 40.6 339.7 1 299.1 1.47 6.0 6.8 None l0 4L9, 332.0 2901 1.52 2-(l-Methyl-5-nitro-2- imidazolyl)-5-piperidinol.3,4-thiadiazole l0 42.0 334.7 292.7 1.48 L0 2.7 None 10 42.4 324.6 282.2 1.53 10 43.7 322.2 278.5 1.53 l-[ 5-( l-Methyl-S-nitro-Z- imidazolyl)- l ,3.4- thiadiazol-Z-yl ]4- l0 42.3 337.8 3 l 5.5 L47 5.0 4.1 (2-tl'1iazolyl )piperazine 10 43.7 346.9 303.2 1.50 9.0 2.0

None 10 40.4 344.6 304.2 1.58 N- {5-1 1- Z-Hydroxyethyl -5- nitro-2-imidazolyl1-l 3.4- thiadiazol-2-yl-acetamide 10 40.4 345.9 305.5 1.57 0.7 0.6

EXAMPLE 55 TABLE XV Basal Swine Ration Ingredient Percent Two experiments are conducted on commerc al Ground yellow com 555 swine farms in Iowa to investigate the effects on daily Soybean oil meal (44% protein) 17.5 gain and feed efficiency of young pigs from feeding var- Rolled Oats Sugar 5.0 lOUS levels of 2-(2-am1no-5-th1ad1azolyl)-l-methyl-S- 50 Dried skim milk 5'0 nitroimidazole in the ration. In each experiment, the ionized whley 2.5 is mea 2.5 pigs are weaned at approximately 4 weeks of age and Dicalcium phospham L5 placed on test immediately. The following expenmen- Iodized salt 0.5 tal treatments are used: f V1tamlns Total 100.0

A. Basal (Unmedicated ration) B. Basal Compound X (50 gm/ton of feed) C. Basal Compound X gm/ton of feed) choline chloride. 130 g.; Vitamin B 20 mg.; folic acid. mg.

TABLE XVI Average Performance Data for Young Pigs Fed Several Levels of 2-(Z-amino-S-thiadiazolyl)-l-methyl-5-nitroimidazaole In The Feed for 42 Days A B D 31 32 TABLE XVI -continued v :Average Performance Data for Young Pigs Fed Several Levels of 2-( 2-amino-5-thiudiazoly'l) 1-methyl-5nitroimiduzuole In The Feed for 42 Days A B. C D E Compound X (gm/ton) AUREO S.P. Control 5 100 200 250 1 Mortality Expt. 1 v 2 r g 0 1 O Expt. 2 3 1 -0 1 1 4 Total '5' i I O 2 I Av. Initial WL, lbs.

Expt. I 14.7 14.7 l4.5 14.9 14.9 Expt..2 17.4 17.0 17.5 17.3 17.5 Average 16.1) 15.8 16.0 16.1 16.2

" AV. DaiIz-Gain, lbs. v I

Ex L-i .56 w .63 '.65. '.71 .69 -;Expt. 2 .51 .60 p .83 a .76 1 .63 V Average I I .54 V 62 .74 I .74 .66

' AvYF'e'ed/Gain Ratio I Expt. 1 '2.4 6 2.43 2.35 a 2.30 2.29 I Expt. 2. g 3.21 2,59 2.40 I 2.49 2.83 Average 2.84 2.51 2.38 2.40 2.56

EXAMPLE 56 EXAMPLE Ingredients 1 Amount mg. Ingredients Amount mg;

2-(Z-umino-S-thiadiazolyhl 480 2-(Z-methyIamino-S-thiadiazblyl)- 60b l-methyl--nitroimidazolc meIhyI-S-nitroimidazole Magnesium stearate 10 Lactose 1000 Lactose 600 Mix thoroughly and place in 6 hard gelatin capsules. Each capsule contains 100 mg. of drug.

We claim: 1

1. A method for enhancing the growth rate of warmbIooded animals which comprises orally-administering to said animals a growth promoting amount of l-[5-( 1'- methyl-5-nitro-2-imidazolyl l ,3,4-thiadiazo1e-2-yl']-4- (2-thiazo1y1)piperazine. i 1 1= After thoroughly mixing the above phase in 12 hard gelatin capsules, each capsule will contain 40 mg. of active drug. 

1. A METHOD FOR ENCHANCING THE GROWTH RATE OF WARMBLOODED ANIMALS WHICH COMPRISES ORALLY ADMINISTERING TO SAID ANIMALS A GROWTH PROMOTING AMOUNT OF 1-(5-(1-METHYL-5NITRO-2-IMIDAZOLYL)-1,3,4-THIADIAZOLE-2-YL)-4-(2 -THIAZOLYL)PIPERAZINE. 